Derivatives of [6,7-dihydro-5H- Imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylic acid amide

ABSTRACT

Derivatives of 6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylic acid amide which exhibit good inhibitory effect upon the interaction of CAMs and Leukointegrins and are thus useful in the treatment of inflammatory disease.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a divisional of U.S. application Ser. No.10/685,638, filed Oct. 15, 2003, which application claims the benefit ofU.S. Provisional Application No. 60/422,449, filed on Oct. 30, 2002,which applications are both herein incorporated by reference in theirentirety.

BACKGROUND OF THE INVENTION

1. Technical Field

The present invention relates generally to a series of novel derivativesof[6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid amide, the synthesis of these compounds and their use in thetreatment of inflammatory disease.

2. Background Information

Research spanning the last decade has helped to elucidate the molecularevents attending cell-cell interactions in the body, especially thoseevents involved in the movement and activation of cells in the immunesystem. See generally, Springer, T. Nature, 1990, 346, 425-434. Cellsurface proteins, and especially the Cellular Adhesion Molecules(“CAMs”) and “Leukointegrins”, including LFA-1, MAC-1 and gp150.95(referred to in WHO nomenclature as CD18/CD11a, CD18/CD11b, andCD18/CD11c, respectively) have correspondingly been the subject ofpharmaceutical research and development having as its goal theintervention in the processes of leukocyte extravasation to sites ofinjury and leukocyte movement to distinct targets. For example, it ispresently believed that prior to the leukocyte extravasation, which is amandatory component of the inflammatory response, activation ofintegrins constitutively expressed on leukocytes occurs and is followedby a tight ligand/receptor interaction between integrins (e.g., LFA-1)and one or several distinct intercellular adhesion molecules (ICAMs)designated ICAM-1, ICAM-2, ICAM-3 or ICAM-4 which are expressed on bloodvessel endothelial cell surfaces and on other leukocytes. Theinteraction of the CAMs with the Leukointegrins is a vital step in thenormal functioning of the immune system. Immune processes such asantigen presentation, T-cell mediated cytotoxicity and leukocyteextravasation all require cellular adhesion mediated by ICAMsinteracting with the Leukointegrins. See generally Kishimoto, T. K.;Rothlein; R. R. Adv. Pharmacol. 1994, 25, 117-138 and Diamond, M.;Springer, T. Current Biology, 1994, 4, 506-532.

A group of individuals has been identified which lack the appropriateexpression of Leukointegrins, a condition termed “Leukocyte AdhesionDeficiency” (Anderson, D. C.; et al., Fed. Proc. 1985, 44, 2671-2677 andAnderson, D. C.; et al., J. Infect. Dis. 1985, 152, 668-689). Theseindividuals are unable to mount a normal inflammatory and/or immuneresponse(s) due to an inability of their cells to adhere to cellularsubstrates. These data show that immune reactions are mitigated whenlymphocytes are unable to adhere in a normal fashion due to the lack offunctional adhesion molecules of the CD 18 family. By virtue of the factthat LAD patients who lack CD 18 cannot mount an inflammatory response,it is believed that antagonism of CD18, CD11/ICAM interactions will alsoinhibit an inflammatory response.

It has been demonstrated that the antagonism of the interaction betweenthe CAMs and the Leukointegrins can be realized by agents directedagainst either component. Specifically, blocking of the CAMs, such asfor example ICAM-1, or the Leukointegrins, such as for example LFA-1, byantibodies directed against either or both of these moleculeseffectively inhibits inflammatory responses. In vitro models ofinflammation and immune response inhibited by antibodies to CAMs orLeukointegrins include antigen or mitogen-induced lymphocyteproliferation, homotypic aggregation of lymphocytes, T-cell mediatedcytolysis and antigen-specific induced tolerance. The relevance of thein vitro studies are supported by in vivo studies with antibodiesdirected against ICAM-1 or LFA-1. For example, antibodies directedagainst LFA-1 can prevent thyroid graft rejection and prolong heartallograft survival in mice (Gorski, A.; Immunology Today, 1994, 15,251-255). Of greater significance, antibodies directed against ICAM-1have shown efficacy in vivo as anti-inflammatory agents in humandiseases such as renal allograft rejection and rheumatoid arthritis(Rothlein, R. R.; Scharschmidt, L., in: Adhesion Molecules; Wegner, C.D., Ed.; 1994, 1-38, Cosimi, C. B.; et al., J. Immunol. 1990, 144,4604-4612 and Kavanaugh, A.; et al., Arthritis Rheum. 1994, 37,992-1004) and antibodies directed against LFA-1 have demonstratedimmunosuppressive effects in bone marrow transplantation and in theprevention of early rejection of renal allografts (Fischer, A.; et al.,Lancet, 1989, 2, 1058-1060 and Le Mauff, B.; et al., Transplantation,1991, 52, 291-295).

It has also been demonstrated that a recombinant soluble form of ICAM-1can act as an inhibitor of the ICAM-1 interaction with LFA-1. SolubleICAM-1 acts as a direct antagonist of CD18,CD11/ICAM-1 interactions oncells and shows inhibitory activity in in vitro models of immuneresponse such as the human mixed lymphocyte response, cytotoxic T cellresponses and T cell proliferation from diabetic patients in response toislet cells (Becker, J. C.; et al, J. Immunol. 1993, 151, 7224 and Roep,B. O.; et al., Lancet, 1994, 343, 1590).

Thus, the prior art has demonstrated that large protein molecules whichantagonize the binding of the CAMs to the Leukointegrins havetherapeutic potential in mitigating inflammatory and immunologicalresponses often associated with the pathogenesis of many autoimmune orinflammatory diseases. However proteins have significant deficiencies astherapeutic agents, including the inability to be delivered orally andpotential immunoreactivity which limits the utility of theses moleculesfor chronic administration. Furthermore, protein-based therapeutics aregenerally expensive to produce.

It follows that small molecules having the similar ability as largeprotein molecules to directly and selectively antagonize the binding ofthe CAMs to the Leukointegrins would make preferable therapeutic agents.

Several small molecules have been described in the literature thataffect the interaction of CAMs and Leukointegrins. For example, U.S.Pat. No. 6,355,664 and the corresponding WO 98/39303 disclose a class ofsmall molecule, having a hydantoin core, that are inhibitors of theinteraction of LFA-1 and ICAM-1. WO 01/07440 A1 discloses compoundshaving this same activity that instead have a6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl core. While thecompounds that are described by WO 01/07440 A1 have a more potentinhibitory affect upon the interaction of CAMs and Leukointegrins thando the hydantoins of U.S. Pat. No. 6,355,664 and the correspondingWO9839303, they nevertheless are not ideal therapeutic agents becausethe rate at which they are metabolized is undesirably high.

Thus, the problem to be solved by the present invention is to find smallmolecules that have not only good inhibitory effect upon the interactionof CAMs and Leukointegrins but that also are metabolized at a rate thatis not overly rapid.

BRIEF SUMMARY OF THE INVENTION

The invention comprises a class of derivatives of[6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid amide and methods for making the same. These compounds are usefulfor the treatment of inflammatory conditions in that they exhibit goodinhibitory effect upon the interaction of CAMs and Leukointegrins andare metabolized fairly slowly. Thus, the invention further comprises theuse of these compounds for the treatment of inflammatory conditions andpharmaceutical compositions comprising the same as active ingredients.

DETAILED DESCRIPTION OF THE INVENTION

In its broadest generic aspect, the invention comprises compounds of theformula I

wherein:

-   R¹ and R² are each, independently selected from the group consisting    of:    -   (A) hydrogen, with the proviso that R¹ and R² are not both        hydrogen atoms;    -   (B) —R¹⁰⁰, which is:    -    a straight or branched alkyl of 1 to 7 carbon atoms or        cycloalkyl of 3 to 6 carbon atoms, which alkyl or cycloalkyl        group is mono- or poly substituted with moieties independently        selected from the group consisting of:        -   (i) oxo,        -   (ii) cyano,        -   (iii) halogen,        -   (iv) moieties of the formula —COOR⁶, wherein R⁶ is a            hydrogen atom, a straight or branched alkyl of 1 to 7 carbon            atoms or cycloalkyl of 3 to 6 carbon atoms,        -   (v) moieties of the formula —OR⁷, wherein R⁷ is a hydrogen            atom, a straight or branched alkyl group of 1 to 7 carbon            atoms or an acyl group of the formula —COR⁸ wherein R⁸ is a            straight or branched alkyl group of 1 to 7 carbon atoms,        -   (vi) moieties of the formula —NR⁹R¹⁰, wherein R⁹ and R¹⁰ are            each, independently selected from the group consisting of:            -   (a) hydrogen,            -   (b) straight or branched alkyl of 1 to 7 carbon atoms,            -   (c) acyl of the formula —COR¹¹ wherein R¹¹ is a straight                or branched alkyl group of 1 to 7 carbon atoms, and            -   (d) groups of the formula —COOR¹² wherein R¹² is a                straight or branched alkyl group of 1 to 7 carbon atoms,        -    or wherein R⁹ and R¹⁰ constitute a bridge consisting of 3-5            methylene groups or 2-4 methylene groups and one oxygen            atom, such that the groups R⁹ and R¹⁰ together with the            nitrogen atom between them form a heterocyclic ring,        -   (vii) saturated, heterocyclic groups, consisting of 3 to 5            methylene groups and one oxygen atom, wherein said            heterocyclic groups are optionally mono- or disubstituted            with moieties that are independently selected from the group            consisting of:            -   (a) oxo and            -   (b) straight or branched alkyl of 1 to 3 carbon atoms;                and        -   (viii) aryl, selected from the class consisting of:            -   (a) furyl,            -   (b) tetrazolyl and            -   (c) thiophenyl;    -   (C) aryl, selected from the group consisting of:        -   (i) biphenyl,        -   (ii) phenyl which is mono- or di-substituted with moieties            independently selected from the group consisting of —NH₂ and            N-morpholino, and        -   (iii) quinolinyl; and    -   (D) unsaturated or partially saturated heterocyclic groups        consisting of 2 to 3 carbon atoms, 1 to 2 nitrogen atoms, 0 to 1        sulfur atoms and 0 to 1 oxygen atoms wherein said heterocyclic        group is optionally mono- or polysubstituted with one or more of        the following moieties independently selected from the group        consisting of:        -   (i) oxo and        -   (ii) straight or branched alkyl of 1 to 7 carbon atoms;-    or wherein R¹ and R² constitute a saturated 3 to 5 methylene group    bridge which together with the nitrogen atom between them form a    heterocyclic ring, wherein said heterocyclic ring is mono- or    disubstituted with moieties independently selected from the group    consisting of:    -   (A) —OH,    -   (B) —COOH and    -   (C) —CONH₂;-   R³ is:    -   (A) aryl selected from the group consisting of pyridyl and        pyrimidyl, wherein one or more hydrogen atoms of said aryl group        are optionally and independently substituted with moieties        selected from the group consisting of:        -   (i) cyano,        -   (ii) halogen and        -   (iii) groups of the formula —NR¹³R¹⁴, wherein R¹³ and R¹⁴            are each, independently, hydrogen or straight or branched            alkyl of 1 to 3 carbon atoms;    -   (B) trifluoromethoxy or,    -   (C) cyano;-   R⁴ is straight or branched alkyl of 1 to 3 carbon atoms;-   R^(5a) is C₁ or CF₃;-   R^(5b) is C₁ or CF₃;-   X is an oxygen or a sulfur atom; and-   Y is an oxygen or a sulfur atom.

In a preferred generic aspect, the invention comprises compounds of theformula I, wherein:

-   R¹ and R² are each independently selected from the group consisting    of:    -   (A) hydrogen with the proviso that R¹ and R² are not both        hydrogen atoms;    -   (B) —R¹⁰⁰, which is:    -    a straight or branched alkyl of 1 to 4 carbon atoms, which        alkyl group is mono- or disubstituted with moieties        independently selected from the group consisting of:        -   (i) oxo,        -   (ii) OH,        -   (iii) moieties of the formula —NR⁹R¹⁰, wherein R⁹ and R¹⁰            are each, independently selected from a group consisting of:            -   (a) hydrogen and            -   (b) methyl,        -   (iv) tetrazole,-    or wherein R¹ and R² constitute a saturated 5 methylene group    bridge which together with the nitrogen atom between them form a    heterocyclic ring, wherein said heterocyclic ring is monosubstituted    with COOH;-   R³ is:    -   (A) aryl selected from the group consisting of 3-pyridyl and        5-pyrimidyl wherein said aryl group is monosubstituted with:        -   (i) cyano or        -   (ii) NH₂,    -   (B) trifluoromethoxy or    -   (C) cyano;-   R⁴ is a methyl group;-   R^(5a) is Cl;-   R^(5b) is Cl;-   X is an oxygen atom and-   Y is an oxygen atom.

In a penultimately preferred generic aspect, the invention comprisescompounds of the formula I wherein:

-   R¹ and R² are each, independently selected from the group consisting    of:    -   (A) hydrogen with the proviso that R¹ and R² are not both        hydrogen atoms, or    -   (B) —R¹⁰⁰, which is:    -    straight or branched alkyl of 1 to 4 carbon atoms, which alkyl        group is mono- or disubstituted with moieties independently        selected from the group consisting of:        -   (i) oxo,        -   (ii) OH and        -   (iii) NH₂;-   R³ is trifluoromethoxy or cyano;-   R⁴ is a methyl group;-   R^(5a) is Cl;-   R^(5b) is Cl;-   X is an oxygen atom; and-   Y is an oxygen atom.

It will be appreciated that the compounds of the formula I have at leasttwo chiral centers. In an ultimately preferred generic aspect, theinvention includes compounds of formula I with the absolutestereochemistry depicted below in formula I*.

Specifically preferred compounds of formula I of the invention are thoseselected from the group consisting of:

-   ({(S)-1-[(R)-5-(4-Cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-acetic    acid,    ({(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-acetic    acid,-   (S)-1-[(R)-5-(4-Cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylic    acid (2-hydroxy-2-methyl-propyl)-amide,-   (S)-1-[(R)-5-(4-Cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylic    acid (2-hydroxy-ethyl)-amide,-   (S)-1-[(R)-5-(4-Cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylic    acid carbamoylmethyl-amide,-   (S)-1-[(R)-5-[4-(2-Cyano-pyridin-3-yl)-benzyl]-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylic    acid (2-hydroxy-2-methyl-propyl)-amide,-   (S)-1-[(R)-5-[4-(2-Cyano-pyridin-3-yl)-benzyl]-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylic    acid carbamoylmethyl-amide,-   (S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylic    acid (1H-tetrazol-5-ylmethyl)-amide,-   (S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-trifluoromethoxy-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylic    acid (2-hydroxy-2-methyl-propyl)-amide, and-   (S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-trifluoromethoxy-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylic    acid carbamoylmethyl-amide.

The invention also includes pharmaceutically acceptable salts of thecompounds of the formula I.

General Synthetic Methods

Compounds of the invention may be prepared by the general methodsdescribed below. Typically, reaction progress may be monitored by thinlayer chromatography (TLC) if desired. If desired, intermediates andproducts may be purified by chromatography on silica gel and/orrecrystallization, and characterized by one or more of the followingtechniques: NMR, mass spectroscopy and melting point. Starting materialsand reagents are either commercially available or may be prepared by oneskilled in the art using methods described in the chemical literature.

Compounds of formula I may be prepared from intermediate II. Thesynthesis of intermediate II is reported by Wu et al., U.S.Non-provisional application Ser. No. 09/604,312 and Frutos et al., U.S.Pat. No. 6,441,183, both incorporated herein by reference.

Intermediate II may be prepared by the procedure illustrated in SchemeI.

As illustrated above, intermediate III is deprotonated with a suitablebase such as lithium bis(trimethylsilyl)amide at about −20° C. to −30°C., and then alkylated with a substituted benzyl halide, preferably abenzyl bromide (IV) to produce V. Hydrolysis of the trifluoroacetamidegroup of V, for example by treatment with 40% aqueousbenzyltrimethylammonium hydroxide in dioxane/50% NaOH, followed bytreatment with acid, such as HCl, provides VI. Treatment of VI withthiocarbonyldiimidazole in the presence of a base such as4-(N,N-dimethylamino)pyridine (DMAP) provides VII. Treatment of VII withaminoacetaldehyde dimethyacetal and t-butylhydroperoxide solution,followed by treatment of the intermediate acetal with an acid such asp-toluenesulfonic acid provides VIII. Iodination of VIII by treatmentwith an iodinating agent such as N-iodosuccinamide provides II.

The method used for preparation of intermediate III, treatment of theamide formed from N-Boc-D-alanine and 3,5-dichloroaniline withtrifluoroacetic acid to remove the Boc-group, followed by treatment withpivalaldehyde, and acylation of the resulting imidazolodone withtrifluoroacetic anhydride is described in U.S. Pat. No. 6,414,161, citedabove, and in the chemical literature (N. Yee, Org Lett., 2000, 2,2781).

The synthesis of compounds of formula I from intermediate II isillustrated in Scheme II.

As illustrated above, treatment of II with a Grignard reagent, such ascyclopentyl magnesium bromide or chloride, followed by treatment of theresulting magnesium salt with SO₂ and then N-chlorosuccinimide providesthe sulfonyl chloride IX. Treatment of 1× with the desired amine (X) inthe presence of a suitable base such as triethylamine, provides thedesired product of formula (I). Intermediates X are either commerciallyavailable or readily prepared from commercially available startingmaterials by methods known in the art. The initial product of formula Imay be further modified by methods known in the art to provideadditional compounds of the invention. Several examples are provided inthe Synthetic Examples section.

The desired R³ on formula I compounds may be obtained by selection ofthe appropriately substituted intermediate IV in Scheme I. Alternately,intermediate VIII having R³ being Br (VIIIa) may be converted tointermediates having R³ being CN or an optionally substituted5-pyrimidyl group as illustrated in Scheme III.

As illustrated above, the aryl bromide VIIIa is treated with a cyanidesalt, preferably CuCN and heated in a suitable solvent such as DMF toprovide the cyano-intermediate VIIb. Treatment of VIIIa with apyrimidine boronate ester such as5-(4,4,5,5,-tetramethyl-[1,3,2]dioxaborolan-2-yl)-pyrimidine in thepresence of a palladium catalyst such as[1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II).CH₂Cl₂(PdCl₂(dppf).CH₂Cl₂) and a base such as potassium carbonate in asuitable solvent (Suzuki reaction), for example dimethoxyethane,provides the pyrimidine intermediate VIIc. Intermediates VIIb and VIIcmay then be converted to desired compounds of formula I by theprocedures described in Schemes I and II. The Suzuki reaction to convertR₃=Br to R₃=an optionally substituted pyrimidine may also be carried outon a compound of formula I.

The invention is further described by the following synthetic examples.

SYNTHETIC EXAMPLES Example 1 Synthesis of(R)-1-(3,5-dichloro-phenyl)-3-methyl-3-(4-trifluoromethoxy-benzyl)-1H-imidazo[1,2-a]imidazol-2-one

Lithium bis(trimethylsilyl)amide (LiHMDS) (38.0 mL, 1 M in THF) wasadded slowly dropwise over 25 min to a solution of(2S,5R)-2-tert-butyl-3-(3,5-dichloro-phenyl)-5-methyl-1-(2,2,2-trifluoro-acetyl)-imidazolidin-4-one(10.0 g, 25.17 mmol) in 60 mL of THF at −20° C. After stirring at −20°C. for 20 min, a solution of 4-trifluoromethoxybenzyl bromide (6.04 mL,37.76 mmol) in 30 mL of THF was added dropwise over 20 min. The mixturewas stirred at −20° C. for 45 min, warmed to −5° C. over 1 h, and thenpoured over 50 mL of ice-cold saturated NH₄Cl solution. The resultingmixture was extracted with two portions of EtOAc (200, 100 mL). Thecombined organic phases were washed with brine, dried over Na₂SO₄,filtered and concentrated. The crude product was triturated with hexanesto afford 12.5 g (87%) of(2R,5R)-2-tert-butyl-3-(3,5-dichloro-phenyl)-5-methyl-1-(2,2,2-trifluoro-acetyl)-5-(4-trifluoromethoxy-benzyl)-imidazolidin-4-oneas an off-white solid.

To a solution of the above imidazolidinone (6.0 g, 10.5 mmol) in 40 mLof dioxane was added 40% aqueous benzyltrimethylammonium hydroxide (6.59g, 15.75 mmol) at room temperature. As the mixture was warmed to 40° C.,50% aqueous sodium hydroxide (1.68 g, 21.0 mmol) was added slowlydropwise over 5 min. The mixture was stirred at 40° C. for 18 h, then asolution of 6.4 g of conc HCl in 3.3 mL of water was added slowlydropwise over 10 min. The mixture was warmed to 50° C. and stirred foran additional 5 h, then cooled to room temperature and concentrated. 50mL of toluene was added to the residue, and the biphasic mixture wasstirred vigorously as 50% aqueous sodium hydroxide (3.0 g) was addedslowly dropwise (pH of the aqueous phase ≧10). The aqueous layer wasextracted with two portions of toluene, and the combined organic phaseswere washed with water and brine, dried over Na₂SO₄, filtered andconcentrated to afford 4.24 g of(R)-2-amino-N-(3,5-dichloro-phenyl)-2-methyl-3-(4-trifluoromethoxy-phenyl)-propionamideas a light brown oil.

To a solution of the above propionamide (4.24 g, 10.41 mmol) in 30 mL ofTHF was added thiocarbonyldiimidazole (2.81 g, 15.77 mmol) and DMAP(0.127 g, 1.04 mmol). The mixture was heated at reflux for 17 h, cooledto room temperature, and concentrated. The orange oily residue wasdissolved in 50 mL of toluene and treated slowly dropwise with 20 mL of5% aqueous HCl solution. After stirring the mixture for 10 min, theaqueous layer was separated and extracted with 30 mL of toluene. Thecombined organic phases were washed with four 20-mL portions of waterand 20 mL of brine, dried over Na₂SO₄, filtered and concentrated toprovide 4.48 g of(R)-3-(3,5-dichloro-phenyl)-5-methyl-2-thioxo-5-(4-trifluoromethoxy-benzyl)-imidazolidin-4-oneas an orange foam.

To a solution of the above thiohydantoin (4.47 g, 9.95 mmol) andaminoacetaldehyde dimethylacetal (6.50 mL, 59.7 mmol) in 20 mL of MeOHwas added 7.69 mL (59.7 mmol, 70% in water) of t-butyl hydroperoxidesolution, dropwise over 25 min. During the addition and for about 1 hafter, the internal temperature of the mixture was kept below 20° C.with an ice water bath. The mixture was stirred at room temperature for86 h, and 25 mL of saturated NaHSO₃ solution was added slowly dropwise,maintaining the internal temperature below 20° C. with an ice waterbath. The resulting cloudy white mixture was concentrated. To theresidue was added EtOAc, and this mixture was concentrated again. Theoily residue was partitioned between 30 mL of EtOAc and 20 mL of water,and the aqueous phase was separated and extracted with 20 mL of EtOAc.The combined organic layers were washed with 25 mL of water and brine,dried over Na₂SO₄, filtered and concentrated to give 5.21 g of(R)-3-(3,5-dichloro-phenyl)-2-[(E)-2,2-dimethoxy-ethylimino]-5-methyl-5-(4-trifluoromethoxy-benzyl)-imidazolidin-4-oneas a thick yellow oil.

A solution of the above crude acetal (5.20 g, 9.95 mmol) in 30 mL ofacetone was treated with p-toluenesulfonic acid (1.89 g, 9.96 mmol). Themixture was heated at reflux for 2 h, then cooled to room temperatureand concentrated. The resulting dark orange oil was dissolved in 40 mLof EtOAc and treated carefully with a solution of 2.3 g of NaHCO₃ in 23mL of water. After gas evolution ceased, the aqueous phase was separatedand extracted with two portions of EtOAc. The combined organic layerswere washed with saturated NaHCO₃ solution, two portions of water, andbrine, dried over Na₂SO₄, filtered and concentrated. The oily residuewas purified by silica gel chromatography to afford 1.58 g of the titlecompound as a thick colorless oil (456.2, M+1).

Example 2 Synthesis of(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-trifluoromethoxy-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonylchloride

A solution of(R)-1-(3,5-dichloro-phenyl)-3-methyl-3-(4-trifluoromethoxy-benzyl)-1H-imidazo[1,2-a]imidazol-2-one(Example 1) (1.54 g, 3.38 mmol) in 30 mL of THF was treated withN-iodosuccinimide (0.846 g, 3.76 mmol) and pyridinium p-toluenesulfonate(0.086 g, 0.37 mmol). The mixture was stirred at room temperature for 17h, then diluted with EtOAc and washed with 10% Na₂S₂O₃ solution andwater. The combined aqueous layers were extracted with 10 mL of EtOAc.The combined organic phases were washed with 25 mL of brine, dried overNa₂SO₄, filtered and concentrated. The crude orange oil was purified bysilica gel chromatography to provide 1.27 g (65%) of(R)-1-(3,5-dichloro-phenyl)-5-iodo-3-methyl-3-(4-trifluoromethoxy-benzyl)-1H-imidazo[1,2-a]imidazol-2-oneas an off-white oil (582.0, M+1).

A solution of the above iodide (1.24 g, 2.13 mmol) in 16 mL of THF wascooled at −40° C. as cyclopentyl magnesium chloride (1.17 mL, 2 M indiethyl ether) was added dropwise over 10 min. After stirring at −40° C.for 1 h, SO₂ (g) was added by placing an inlet needle just above thesurface of the reaction mixture for 1.5 min. The bright yellow mixturewas warmed to −20° C. over 1 h and then stirred at room temperature for1 h. N₂ (g) was bubbled through the mixture for 20 min followed byconcentration and pumping under high vacuum for 12 h. The resultingyellow foam was dissolved in 16 mL of THF and cooled at −20° C. as asolution of N-chlorosuccinimide (0.341 g, 2.56 mmol) in 8 mL of THF wasadded dropwise over 5 min. After stirring at −20° C. for 1 h, themixture was poured over ice and extracted with two portions of EtOAc.The combined organic layers were washed with 20 mL of ice-cold brine,dried over Na₂SO₄, filtered and concentrated. Purification by silica gelchromatography provided 0.975 g (83%) of the title compound as a thickoil (554.2, M+1).

Example 3 Synthesis of(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-trifluoromethoxy-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-hydroxy-2-methyl-propyl)-amide

To a suspension of(S)-2-(2-hydroxy-2-methyl-propylcarbamoyl)-pyrrolidine-1-carboxylic acidtert-butyl ester (see Example 11) (0.190 g, 0.663 mmol) in 1 mL ofdioxane was added HCl (2.0 mL, 4 M in dioxane), and the resulting cloudymixture was stirred at room temperature for 4 h. Concentration of themixture was followed by addition of CH₂Cl₂, and this process wasrepeated twice. Final pumping under high vacuum for 12 h afforded thedeprotected amine HCl salt as a pale yellow oil. This crude amine HClsalt was dissolved in 3 mL of CH₂Cl₂ and treated with triethylamine(0.200 mL, 1.44 mmol). After stirring at room temperature for 10 min, asolution of(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-trifluoromethoxy-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonylchloride (Example 2) (0.199 g, 0.359 mmol) in 3 mL of CH₂Cl₂ was addedrapidly dropwise via cannula. The reaction mixture was stirred at roomtemperature for 3 h, then partitioned between 30 mL of CH₂Cl₂ and 10 mLof water. The organic phase was washed with 10 mL of brine, dried overNa₂SO₄, filtered and concentrated. The crude product was purified bysilica gel chromatography to give 0.228 g (90%) of the title compound asa white foam (704.0, M+1).

Example 4 Synthesis of(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-trifluoromethoxy-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-hydroxy-ethyl)-amide

To a suspension of(S)-2-(2-hydroxy-ethylcarbamoyl)-pyrrolidine-1-carboxylic acidtert-butyl ester (0.200 g, 0.774 mmol) in 1 mL of dioxane was added HCl(2.0 mL, 4 M in dioxane), and the resulting cloudy mixture was stirredat room temperature for 4 h. Concentration of the mixture was followedby addition of CH₂Cl₂. This process was repeated twice and final pumpingunder high vacuum for 12 h afforded the deprotected amine HCl salt as awhite oil. This crude amine HCl salt was dissolved in 3 mL of DMF andtreated with triethylamine (0.212 μL, 1.52 mmol). After stirring at roomtemperature for 10 min, a solution of(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-trifluoromethoxy-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonylchloride (Example 2) (0.211 g, 0.380 mmol) in 4 mL of CH₂Cl₂ was addedrapidly dropwise via cannula. The reaction mixture was stirred at roomtemperature for 2 h. Following the addition of 75 mL of EtOAc, theorganic layer was washed with three portions of water, then brine, driedover Na₂SO₄, filtered and concentrated. The crude product was purifiedby preparative TLC to afford 0.218 g (85%) of the title compound as awhite foam (676.1, M+1).

The following compound was prepared by a procedure analogous to thatdescribed above in Example 4:

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-trifluoromethoxy-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid carbamoylmethyl-amide. (689.3, M+1)

Example 5 Synthesis of(R)-2-({(S)-1-[(R)-5-(4-cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-propionicacid

(R)-3-(4-Bromobenzyl)-1-(3,5-dichlorophenyl)-3-methyl-imidazo[1,2-a]-imidazol-2-one(13.5 g, 29.9 mmol) was dissolved in anhydrous DMF (70 mL) and CuCN(3.22 g, 35.9 mmol) was added. The reaction mixture was heated to 140°C. under argon for 40 h. When the reaction was complete, the solvent wasremoved under reduced pressure and the residue was diluted with CH₂Cl₂.The organic solution was washed four times alternately with 5% aqueouspyridine then water. The organic layer was dried over Na₂SO₄ andfiltered through a diatomaceous earth pad. The filtrate was concentratedand purified by silica gel column chromatography using a mixture ofCH₂Cl₂-hexane (10:1) as an eluent to afford 9.19 g of(R)-3-(4-cyanobenzyl)-1-(3,5-dichlorophenyl)-3-methyl-imidazo[1,2-a]-imidazol-2-one.

To a solution of the above product (5 g, 12.6 mmol) in dry CH₂Cl₂ (50mL) was added pyridinium p-toluenesulfonate (PPTS, 0.32 g, 1.26 mmol)and then N-iodosuccinimide (NIS, 3.39 g, 15.1 mmol) was added in smallportions over a period of 30 min at −20° C. The reaction mixture wasstirred at room temperature overnight covered with aluminum foil. Thereaction mixture was washed with 10% aqueous sodium bisulfite thenwater. The organic phase was dried over Na₂SO₄ and concentrated. Thecrude product was purified by silica gel column chromatography using amixture of hexane-EtOAc (4:1) to afford 5.2 g of(R)-3-(4-cyanobenzyl)-1-(3,5-dichlorophenyl)-5-iodo-3-methyl-1-H-imidazo[1,2-a]-imidazol-2-one.

To a solution of above the above iodide (5.3 g, 10.1 mmol) in anhydrousTHF (50 mL) at −40° C., cyclopentylmagnesium bromide (2M in ether, 6.1mL, 12.2 mmol) was slowly added. The mixture was stirred at −40° C. for30 min and SO₂ was bubbled into the solution for 1 min. The reactionmixture was stirred at −40° C. for 30 min, and then at room temperaturefor another 30 min. The solvent was removed under reduced pressure andthe resulting residue was dried in vacuo for 2 h. The resultingmagnesium sulfonate was dissolved in anhydrous THF and cooled down to−30° C. N-Chlorosuccinimide (NCS, 2.0 g, 15.2 mmol) was added to thesolution and the reaction was stirred at −20° C. for 1 h. The reactionmixture was poured into ice water and extracted with EtOAc. The organicphase was dried over Na₂SO₄ and concentrated. Silica gel columnchromatography using a mixture of hexane-EtOAc (5:1) as an eluentafforded 3.0 g of sulfonyl chloride.

The above sulfonyl chloride (1.0 g, 2.02 mmol) was dissolved in CH₂Cl₂(10 mL) and L-proline t-butyl ester (1.03 g, 6.06 mmol) was added to thesolution. The reaction mixture was stirred at room temperature for 1 hand diluted with CH₂Cl₂ (5 mL). The organic solution was washed with 1%HCl, saturated NaHCO₃ and water. The organic phase was dried over Na₂SO₄and concentrated. Silica gel column chromatography using a mixture ofhexane-EtOAc (1:1) as an eluent afforded 0.56 g of the desired product.This product was then treated with 50% trifluoroacetic acid (TFA) inCH₂Cl₂ at room temperature for 2 h. The reaction mixture was dilutedwith CH₂Cl₂ and washed with water. The organic layer was dried overNa₂SO₄ and concentrated to afford 0.45 g of the desired carboxylic acid.

The above carboxylic acid (0.15 g, 0.26 mmol) was dissolved in a mixtureof CH₂Cl₂-DMF (5 mL-0.1 mL). To this solution, were added D-alaninet-butyl ester (0.07 g, 0.39 mmol) andO-benzotriazole-1-yl-N,N,N′,N′-tetramethyluronium tetrafluoroborate(TBTU, 0.13 g, 0.39 mmol), followed by N,N-diisopropylethylamine (DIPEA,0.11 mL, 0.65 mmol). The reaction mixture was stirred at roomtemperature for 2 h and diluted with CH₂Cl₂. The solution was washedwith 1N HCl and saturated NaHCO₃. The organic phase was then dried overNa₂SO₄ and concentrated. The crude product was purified by preparativethin layer chromatography using EtOAc as an eluent to afford the desiredproduct. The resulting compound was then treated with 50% TFA in CH₂Cl₂at room temperature for 2 h. The reaction mixture was diluted withCH₂Cl₂ and washed with water. The organic layer was dried over Na₂SO₄and concentrated. The resulting compound was purified by preparativethin layer chromatography using CH₂Cl₂-MeOH (95:5) as an eluent toafford 0.119 g of the title compound as a white foam (M+1, 645.2).

The following compound was made by procedures analogous to thosedescribed in the above example:

(S)-1-[(R)-5-(4-cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-aceticacid (631.2, M+1)

Example 6 Synthesis of1-{(S)-1-[(R)-5-(4-cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-piperidine-4-carboxylicacid

To a stirred solution of(S)-1-[(R)-5-(4-cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (see Example 5) (1 eq) in anhydrous CH₂Cl₂ andO-(7-azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (HATU) (1.5 eq) was added DIPEA (3 eq) and themixture was stirred at room temperature for 10 min under nitrogen.Methylisonipecotate (1.5 eq) was added to the reaction mixture andstirred for 2 h at room temperature under nitrogen. The material waspurified by column chromatography over silica gel (gradient 10-25% EtOAcin hexane) to provide the methyl ester. The resulting ester (0.140 g,0.2 mmol) was dissolved in hydrobromic acid in acetic acid (1.0 M) andthe reaction was stirred for 4 h. The mixture was concentrated, toluenewas added, and the mixture was concentrated again. The resulting residuewas chromatographed over silica gel (10% MeOH in methylene chloride) toprovide 12 mg of the title compound as a white solid (M+1, 685).

Example 7(S)-1-[(R)-5-(4-cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid ((S)-2-hydroxy-propyl)-amide

To a stirred solution of(S)-1-[(R)-5-(4-cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (see Example 5) (82 mg, 0.14 mmol) in anhydrous methylene chloride(1 mL) and HATU (82 mg, 0.21 mmol) was added DIPEA (73 μL, 0.42 mmol)and the mixture was stirred for 10 min at room temperature undernitrogen. (S)-1-Amino-propan-2-ol (0.21 mmol) was added and the mixturewas stirred for 2 h at room temperature under nitrogen. The material waspurified by column chromatography over silica gel to provide 62 mg ofthe title compound as a white solid (M+1, 631.2).

The following compounds were made by procedures analogous to thosedescribed in the above example:

(S)-1-[(R)-5-(4-Cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-hydroxy-1,1-dimethyl-ethyl)-amide

(M+1, 645.1);(S)-1-[(R)-5-(4-Cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (furan-2-ylmethyl)-amide

(M+1, 653.2);

Example 8 Synthesis of(S)-1-[(R)-5-(4-cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (4-hydroxy-phenyl)-amide

(S)-1-[(R)-5-(4-Cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (see Example 5) (122 mg, 0.2 mmol) was dissolved in methylenechloride (2 mL) under a nitrogen atmosphere. HATU (121 mg, 0.3 mmol) andDIPEA (111 μL, 0.64 mmol) were added. The t-butyldimethylsilyl-(TBDMS-)protected aniline (71 mg, 0.3 mmol) was added and the reaction wasstirred overnight. The solvent was evaporated and the resulting residuechromatographed over silica gel (gradient EtOAc/hexanes) to provide awhite solid. The white solid was dissolved in methylene chloride (2 mL)and a 1.0 M solution in THF of tetrabutylammonium fluoride (TBAF) (1 mL)was added and the reaction stirred for 3 h. Water and a few drops of 1 NHCl were added and the layers separated. The organic layer was driedover sodium sulfate, filtered, and evaporated. The resulting residue waschromatographed over silica gel (25% EtOAc in methylene chloride) toprovide 83 mg of the title compound as a white solid (M+1, 665.2).

The following compound was made by procedures analogous to thosedescribed in the above example:

(S)-1-[(R)-5-(4-Cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (3-hydroxy-phenyl)-amide

(M+1, 665).

Example 9 Synthesis of(S)-1-[(R)-5-(4-cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid acetyl-amide

To a stirred solution of(R)-5-(4-cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonylchloride (see Example 5) (50 mg, 0.10 mmol) in anhydrous methylenechloride (1 mL) was added 4-dimethylaminopyridine (DMAP) (37 mg, 0.30mmol). L-Proline amide hydrochloride (69 mg, 0.30 mmol) was added to themixture and the reaction was stirred for 18 h at room temperature undernitrogen. The reaction was concentrated and the residue was purified bycolumn chromatography over silica gel (gradient elution with 35-50%EtOAc in hexanes) to provide 141 mg of(S)-1-[(R)-5-(4-Cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid amide as a white solid.

(S)-1-[(R)-5-(4-Cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid amide (98 mg, 0.17 mmol) was dissolved in acetic anhydride (3 mL,0.05 M). The reaction was heated to 100° C. for 18 h. The mixture wasconcentrated, toluene added, and the mixture concentrated again. Theresidue was chromatographed over silica with a solvent system of 10%MeOH in methylene chloride to provide 19 mg of the title compound as awhite solid (M+1, 615).

Example 10 Synthesis of(S)-1-[(R)-5-(4-cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid ((R)-5-oxo-tetrahydro-furan-2-ylmethyl)-amide

(R)-(−)-Dihydro-5-(hydroxymethyl)-2(3H)-furanone (2.0 g, 0.017 mol) wasdissolved in CHCl₃ (20 mL) and cooled to 0° C. Pyridine (4.77 g, 0.060mol) was added to the solution and stirred for 15 min. To this reactionsolution, was added 4-nitrobenzenesulfonyl chloride (Nos-Cl) (4.6 g,0.021 mol) and the reaction mixture was stirred at 0° C. for another 3h. The reaction mixture was then washed with 1N HCl, saturated NaHCO₃and water. The organic phase was dried over Na₂SO₄ and concentrated toafford the Nos-protected intermediate. This intermediate wascharacterized by ¹H-NMR and then used directly for the next reaction. Itwas dissolved in MeOH (20 mL) and NaN₃ (5.6 g, 0.086 mol) was added tothe solution. This heterogeneous reaction mixture was heated to 62° C.and stirred overnight. TLC indicated the reaction was not complete sothe reaction mixture was stirred at 50° C. for 3 more days. The reactionmixture was then concentrated and the resulting residue was diluted withCH₂Cl₂, then washed with water to remove any remaining NaN₃. The organicphase was dried over Na₂SO₄ and concentrated. The desired(R)-(−)-dihydro-5-(azidoymethyl)-2(3H)-furanone was isolated by silicagel column chromatography using CH₂Cl₂ as an eluent.

The above intermediate (0.43 g) was dissolved in MeOH and Pd/C was addedto the solution. The reaction mixture was saturated with H₂ and 4N HClin dioxane was added to the mixture. The reaction mixture was stirred atroom temperature for 2 h and then filtered through a diatomaceous earthpad. The filtrate was concentrated under reduced pressure to afford 0.4g of a mixture of two products (furanone and hydroxy-ester) inhydrochloride salt form.

To a solution of(S)-1-[(R)-5-(4-cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (see Example 5) (0.1 g, 0.174 mmol) in anhydrous DMF was added theabove mixture of amines (0.030 g), followed by TBTU (0.084 g, 0.261mmol) followed by DIPEA (0.075 mL, 0.435 mmol). The reaction mixture wasstirred at room temperature for 30 min. The reaction mixture was thendiluted with EtOAc and washed with water (×3), 1N HCl, and saturatedNaHCO₃. The organic phase was dried over Na₂SO₄ and concentrated. Theresulting residue was dissolved in CH₂Cl₂ and trifluoroacetic acid (1eq) was added to the solution. The reaction mixture was stirred at roomtemperature for 3 h and washed with saturated NaHCO₃ and water. Theorganic phase was dried over Na₂SO₄ and concentrated. The resultingresidue was purified by silica gel preparative thin layer chromatographyusing CH₂Cl₂-MeOH (95:5) as an eluent to afford 0.063 g of the titlecompound as a white foam (M+1, 671.1).

Example 11 Synthesis of(S)-1-[(R)-5-(4-cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-hydroxy-2-methyl-propyl)-amide

A 500 mL 3-necked round bottom flask was fitted with an overheadstirrer, a thermoprobe, and a Claisen head topped with a meteringaddition funnel and N₂ inlet. Lithium aluminum hydride (8.4 g) wasplaced in a flask and the flask was cooled in an ice bath. THF (150 mL)was added to the flask under N₂ stream. Gas was evolved and the internaltemperature rose to ˜50° C. The addition funnel was charged with 10 mLof acetone cyanohydrin in 50 mL of THF. After the internal temperaturecooled to about 5° C., acetone cyanohydrin was added. The rate ofaddition was set to maintain the temperature below 10° C. The reactionmixture was allowed to gradually warm to room temperature and stirovernight. The reaction mixture was then cooled to about 5° C. andNa₂SO₄* 110H₂O was added in portions to maintain the temperature atabout 10° C. After gas evolution and temperature increase stopped, theremaining amount was added and stirred in the ice bath for 30 min. Theice bath was removed and stirring continued overnight. The reactionmixture was filtered and the salts were washed with 200 mL of warm THF(about 50° C.). The filtrate was combined and concentrated to afford 5.6g of 2-hydroxy-2-methyl-propyl-1-amine.

A solution mixture of L-Boc-proline (1.5 g, 7 mmol),2-hydroxy-2-methyl-propyl-1-amine (1.24 g, 13.9 mmol) and HOBt (0.96 g,7.1 mmol) in anhydrous acetonitrile (35 mL) was cooled down to 0° C. andEDC (1.6 g, 8.4 mmol) was added to the reaction mixture in one portion.The reaction mixture was allowed to warm to room temperature and stirredovernight. The solvent was removed, diluted with CH₂Cl₂ (30 mL) andwashed with 5% citric acid (4×5 mL), saturated NaHCO₃ (4×5 mL) andbrine. The organic layer was dried over Mg₂SO₄ and concentrated toafford 1.5 g of the amide.

The Boc-protected alcohol (0.31 g, 1.082 mmol) from above was dissolvedin CH₂Cl₂ (5 mL) and 4N HCl in dioxane (5 mL) was added to the reactionsolution. The reaction mixture was stirred at room temperature for 3 hand then concentrated under reduced pressure to afford 0.24 g ofpyrrolidine-2-carboxylic acid (2-hydroxy-2-methyl-propyl)-amidehydrochloride.

Pyrrolidine-2-carboxylic acid (2-hydroxy-2-methyl-propyl)-amidehydrochloride (0.241 g, 1.082 mmol) was dissolved in CH₂Cl₂ andtriethylamine (0.206 mL, 1.513 mmol) was added to the solution. Thereaction mixture was stirred at room temperature for 10 min.(R)-5-(4-Cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonylchloride (see Example 5) (0.3 g, 0.605 mmol) in CH₂Cl₂ was added to thereaction mixture and stirred for 1 h. DMF (0.5 mL) was then added to thereaction mixture and stirred for another 30 min. The reaction solutionwas washed with 1N HCl and saturated NaHCO₃. The organic phase was driedover Na₂SO₄ and concentrated. The crude product was purified by silicagel preparative thin layer chromatography using CH₂Cl₂-MeOH (10:1) as aneluent to afford 0.28 g of the title compound (M+1, 644.98).

Example 12 Synthesis of(S)-1-[(R)-5-(4-cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid carbamoylmethyl-amide

(S)-1-[(R)-5-(4-Cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (see Example 5) (0.25 g, 0.435 mmol) was dissolved in a mixture ofCH₂Cl₂ (10 mL) and DMF (0.2 mL). To this solution, were addedglycinamide hydrochloride (0.072 g, 0.653 mmol), TBTU (0.21 g, 0.653mmol), and DIPEA (0.19 mL, 1.087 mmol). The reaction mixture was stirredat room temperature for 1 h. The reaction mixture was diluted withCH₂Cl₂ and washed with 1N HCl followed by saturated aq. NaHCO₃. Theorganic phase was dried over Na₂SO₄ and concentrated. The crude productwas purified by silica gel preparative thin layer chromatography usingCH₂Cl₂-MeOH (95:5) as an eluent to afford 0.155 g of the title compoundas a white foam (M+1, 630).

Example 13 Synthesis of(S)-1-[(R)-5-(4-cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-hydroxy-ethyl)-amide

A mixture of L-Boc-proline (2.5 g, 11.6 mmol), HOBt (1.6 g, 11.8 mol)and ethanolamine (0.85 g, 13.9 mmol) in CH₃CN was cooled to 0° C. Tothis heterogeneous reaction mixture, was added EDC (2.67 g, 13.9 mmol)in one portion. The reaction mixture was allowed to warm to roomtemperature and stirred overnight. The solvent was then evaporated andthe residue was treated with EtOAc (100 mL). This organic solution waswashed with 5% citric acid. The aqueous layer was saturated with NaCland extracted with EtOAc (4×50 mL). The organic phase was combined andwashed with saturated NaHCO₃ (10 mL), brine and dried over Na₂SO₄. Thesolution was concentrated and re-dissolved in CH₂Cl₂ (100 mL) and washedwith saturated NaHCO₃ (10 mL) to remove residual HOBt. The organic phasewas dried over Na₂SO₄ and concentrated to afford 2.53 g of theethanolamine coupled proline derivative as a white solid.

The above intermediate (2.53 g) was dissolved in CH₂Cl₂ (30 mL) and 4NHCl in dioxane (7.3 mL) was added to the solution. The reaction solutionwas stirred at room temperature for 2 h. The reaction solution wasbubbled with a strong stream of N₂ for 40 min and then concentrated invacuo to afford the de-protected proline amine HCl salt as a solidifiedgummy residue.

The above amine HCl salt (0.93 g) was dissolved in anhydrous DMF (9 mL)by vigorously stirring at room temperature for 30 min. When all theamine salt was dissolved, triethylamine (1 mL) was added to thesolution.(R)-5-(4-Cyano-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonylchloride (see Example 5) (1.18 g) in anhydrous DMF (1 mL) was then addedto the reaction mixture and stirred at room temperature for 30 min. Thereaction mixture was then diluted with EtOAc and washed with water (×2),1N HCl, saturated NaHCO₃ and brine. The organic phase was then driedover Na₂SO₄ and concentrated. The residue was purified by silica gelcolumn chromatography using CH₂Cl₂ and MeOH (gradient 0-5% MeOH) as aneluent to afford 1.18 g of the title compound as a white foam (M+1,617.2)

Example 14 Synthesis of(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid

Cyclopentylmagnesium bromide (5.2 mL, 2 M solution in Et₂O) was added toa solution of(R)-3-(4-bromo-benzyl)-1-(3,5-dichloro-phenyl)-5-iodo-3-methyl-1H-imidazo[1,2-a]imidazol-2-one(5.0 g) in Et₂O (30 mL) at −40° C. and stirred for 15 min. SO₂ wasbubbled through the reaction solution for 1 min and the reaction mixturewas stirred for an additional 20 min then warmed to room temperature.The precipitate was isolated by filtration, washed with Et₂O and theremainder of the solvent was removed in vacuo. The resultant salt wasdissolved in THF (10 mL) and added dropwise to a solution ofN-chlorosuccinimide (0.86 g) in THF (20 mL) at −25° C. The reaction wasallowed to slowly warm to room temperature and stirred for 1 h.L-Proline-tert-butyl ester (2.2 g) was added to the reaction solutionand stirred for 2 h. The volatiles were removed and the resultantresidue was re-dissolved in EtOAc and washed with 1 N HCl, followed bysaturated NaHCO₃ and H₂O. The organic layers were combined, dried(Na₂SO₄), filtered and concentrated. The resultant residue was purifiedby silica gel chromatography to afford(S)-1-[(R)-5-(4-bromo-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid tert-butyl ester.

To the above tert-butyl ester (2.0 g) in degassed dimethoxyethane (30mL) was added5-(4,4,5,5,-tetramethyl-[1,3,2]dioxaborolan-2-yl)-pyrimidine (1.2 g),K₂CO₃ (1.61 g), and PdCl₂(dppf)-CH₂Cl₂ (0.24 g). The reaction was heatedto 95° C. for 24 h then cooled to room temperature and the volatileswere removed. The residue was diluted in CH₂Cl₂ and washed with H₂O. Theorganic layers were combined, dried (Na₂SO₄), filtered and concentrated.The resultant residue was purified by silica gel chromatography toafford(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid tert-butyl ester (1.5 g).

The above intermediate was dissolved in 50% trifluoroacetic acid-CH₂Cl₂(30 mL) at 0° C. and allowed to slowly warm to room temperature. Thereaction was allowed to stir for 1 h then concentrated to afford 1.2 gof the title compound.

Example 15 Synthesis of(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-methoxy-ethyl)-amide

TBTU (0.077 g), DIPEA (0.090 mL) and(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (Example 14) (0.10 g) were combined in 5% DMF-CH₂Cl₂ (4.2 mL) atroom temperature. 2-Methoxyethylamine (0.036 g) was then added and thereaction solution was stirred overnight. The reaction was diluted withCH₂Cl₂, poured into 1N HCl, and extracted with CH₂Cl₂. The organiclayers were subsequently extracted with CH₂Cl₂ from saturated aqueousNaHCO₃ followed by brine. The combined organic phase was dried (MgSO₄),filtered, and concentrated. The residue was purified by silica gelchromatography to afford 0.045 g of the title compound (684.2, M+1) as afoam.

Analogous procedures were employed to prepare the following compoundsand utilized standard coupling reagents, such as TBTU, carbonyldiimidazole (CDI), or N-cyclohexylcarbodiimide; tertiary amines, such asEt₃N or DIPEA; and either the amine or amine hydrochloride:

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-acetylamino-ethyl)-amide (711.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-dimethylamino-ethyl)-amide (697.3, M+1)

Acetic acid2-({(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-ethylester (712.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-hydroxy-2-methyl-propyl)-amide (698.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-hydroxy-ethyl)-amide

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-morpholin-4-yl-ethyl)-amide (739.3, M+1)

(R)-1-(3,5-Dichloro-phenyl)-5-[(S)-2-(3-hydroxy-piperidine-1-carbonyl)-pyrrolidine-1-sulfonyl]-3-methyl-3-(4-pyrimidin-5-yl-benzyl)-1H-imidazo[1,2-a]imidazol-2-one(710.3, M+1)

[2-({(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-ethyl]-carbamicacid tert-butyl ester (769.3, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-amino-ethyl)-amide (669.3, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (3-hydroxy-propyl)-amide (684.3, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (furan-2-ylmethyl)-amide (706.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2,3-dihydroxy-propyl)-amide (700.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-hydroxy-1-methyl-ethyl)-amide (684.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid cyanomethyl-amide (665.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid ((R)-2-hydroxy-1-methyl-ethyl)-amide (684.3, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid ((S)-1-hydroxymethyl-3-methyl-butyl)-amide (726.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid ((R)-1-hydroxymethyl-3-methyl-butyl)-amide (726.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-hydroxy-1-hydroxymethyl-ethyl)-amide (700.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-amino-phenyl)-amide (717.2, M+1)

(S)-1′-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (3-amino-phenyl)-amide (717.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (4-amino-phenyl)-amide (717.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid biphenyl-4-ylamide (778.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid quinolin-6-ylamide (753.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (4-morpholin-4-yl-phenyl)-amide (787.2, M+1)

(S)-1′-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (1-methyl-4-oxo-4,5-dihydro-1H-imidazol-2-yl)-amide (722.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (1,3,5-trimethyl-1H-pyrazol-4-yl)-amide (734.3, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (4-oxo-4,5-dihydro-thiazol-2-yl)-amide (725.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid [1,3,4]thiadiazol-2-ylamide (710.1, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-ethyl-2H-pyrazol-3-yl)-amide (720.3, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-hydroxy-1,1-dimethyl-ethyl)-amide (698.1, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid ((S)-2-hydroxy-propyl)-amide (684.0, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid ((R)-2-hydroxy-propyl)-amide (684.0, M+1)

(R)-1-(3,5-Dichloro-phenyl)-5-[(S)-2-((R)-3-hydroxy-pyrrolidine-1-carbonyl)-pyrrolidine-1-sulfonyl]-3-methyl-3-(4-pyrimidin-5-yl-benzyl)-1H-imidazo[1,2-a]imidazol-2-one(696.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid methylcarbamoylmethyl-amide (697.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid ((S)-1-methylcarbamoyl-ethyl)-amide (711.2, M+1)

1-{(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-piperidine-4-carboxylicacid amide (737.3, M+1)

(R)-1-(3,5-Dichloro-phenyl)-5-[(S)-2-((S)-3-hydroxy-pyrrolidine-1-carbonyl)-pyrrolidine-1-sulfonyl]-3-methyl-3-(4-pyrimidin-5-yl-benzyl)-1H-imidazo[1,2-a]imidazol-2-one(696.2, M+1)

1-({(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-cyclopropanecarboxylicacid methyl ester (724.2, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (4,5-dihydro-oxazol-2-yl)-amide (695.0, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (1H-tetrazol-5-ylmethyl)-amide (708.0, M+1)

Example 16 Synthesis of(R)-2-({(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-propionicacid

TBTU (0.078 g) and(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (Example 14) (0.10 g) were combined in 7% DMF-CH₂Cl₂ (3.2 mL) atroom temperature. D-Alanine-tert-butyl ester hydrochloride (0.044 g),followed by DIPEA (0.07 mL), was then added and the reaction solutionwas stirred for 16 h. The reaction was diluted with CH₂Cl₂, poured into1N HCl, and extracted with CH₂Cl₂. The organic layers were washed withsaturated aqueous NaHCO₃ followed by H₂O. The combined organic phase wasdried (MgSO₄), filtered and concentrated. The resultant residue wasre-dissolved in either 50% trifluoroacetic acid-CH₂Cl₂ or 4N HCl-dioxane(5 mL) and stirred at room temperature for 2 h. Following aqueous workupthe residue was purified by silica gel chromatography to afford thetitle compound (0.045 g) as a foam (698.9, M+1).

Analogous procedures were employed to prepare the following compoundsand utilized standard coupling reagents, such as TBTU, CDI, orN-cyclohexylcarbodiimide, and tertiary amines, such as Et₃N or DIPEA:

(S)-2-({(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-propionicacid (698.3, M+1)

({(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-aceticacid (683.9, M+1)

({(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-methyl-amino)-aceticacid (698.1, M+1)

2-({(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-2-methyl-propionicacid (712.1, M+1)

3-({(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-propionicacid (698.0, M+1)

Example 17 Synthesis of1-{(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-piperidine-4-carboxylicacid

TBTU (0.12 g),(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (Example 14) (0.15 g) and methyl isonipecotate (0.041 g), followedby DIPEA (0.10 mL), were combined in 1% DMF-CH₂Cl₂ (10.1 mL) at roomtemperature and the solution was stirred for 1 h. The reaction wasdiluted with CH₂Cl₂, poured into 1N HCl, and extracted with CH₂Cl₂. Thecombined organic layers were washed with saturated aqueous NaHCO₃followed by H₂O. The organic phase was dried (Na₂SO₄), filtered andconcentrated. The resultant residue was re-dissolved in 30% HBr—AcOH andheated to 95° C. for 3 h. Following aqueous workup, the organic layerwas dried (Na₂SO₄), filtered, concentrated, and the residue was purifiedby silica gel chromatography to afford the title compound (0.099 g)(738.2, M+1).

Analogous procedures were employed to prepare the following compoundsand utilized standard coupling reagents, such as TBTU, CDI, orN-cyclohexylcarbodiimide; tertiary amines, such as Et₃N or DIPEA; and anamino acid as either the methyl or ethyl ester.

3-({(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-4,4,4-trifluoro-butyricacid (766.3, M+1)

(S)-2-({(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-methyl-amino)-3-methyl-butyricacid (740.3, M+1)

(1S,2S)-2-({(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-cyclohexanecarboxylicacid (752.2, M+1)

3-({(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-butyricacid (712.1, M+1);

3-({(S)-1-[(R)-7-(3,5-Dichloro-phenyl)—methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-2-methyl-propionicacid (712.0, M+1)

1-({(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-cyclopropanecarboxylicacid (710.1, M+1)

Example 18 Synthesis of(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-carbamoyl-ethyl)-amide.

TBTU (0.15 g),(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (Example 14) (0.20 g), β-alanine methyl ester (0.067 g) and DIPEA(0.14 mL) were combined in 1% DMF-CH₂Cl₂ (10.1 mL) at room temperatureand the solution was stirred for 1 h. The reaction was diluted withCH₂Cl₂, poured into 1N HCl, and extracted with CH₂Cl₂. The organiclayers were combined and washed with saturated aqueous NaHCO₃ followedby H₂O. The combined organic phase was dried (Na₂SO₄), filtered andconcentrated. The resultant residue was re-dissolved in 2N NH₃—CH₃OH (10mL) and heated to 40-60° C. for 48 h. The volatiles were removed and theresidue was purified by silica gel chromatography to afford the titlecompound (0.021 g) as a foam (697.2, M+1).

Example 19 Synthesis of(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid ((R)-1-carbamoyl-ethyl)-amide

N-Hydroxysuccinimide (0.035 g), followed by 1,3-dicyclohexylcarbodiimide(DCC, 0.064 g), were added to a solution of(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (Example 14) (0.16 g) in CH₂Cl₂ (20 mL). The reaction was stirredat room temperature for 1 h, then was filtered through a pad ofdiatomaceous earth, concentrated and re-dissolved in 2N NH₃—CH₃OH (10mL). The solution was stirred at room temperature for 1 h. The volatileswere then removed and the resultant residue was re-dissolved in CH₂Cl₂and washed with H₂O. The combined organic layers were concentrated andthe resultant residue was purified by silica gel chromatography toafford the title compound (0.128 g) as a foam (697.2, M+1).

The following compounds were prepared by procedures analogous to thosedescribed in the above example:

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid carbamoylmethyl-methyl-amide (697.3, M+1)

(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (1-carbamoyl-1-methyl-ethyl)-amide (711.2, M+1)

Example 20 Synthesis of(S)-2-({(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-3-hydroxy-propionicacid.

TBTU (0.077 g), DIPEA (0.1 ml),(S)-1-[(R)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (Example 14) (0.075 g) and O-tert-butyl-L-serine tert-butyl ester(0.061 g) were combined in 1:4 DMF-CH₂Cl₂ (1.2 mL) at room temperatureand stirred for 2 h. The reaction was diluted with CH₂Cl₂ and extractedwith 1 N HCl, followed by saturated aqueous NaHCO₃ and brine. Thecombined organic layers were dried (MgSO₄), filtered and concentrated.The resultant residue was purified by silica gel chromatography toafford the desired ester (0.099 g).

The above ester (0.099 g) was dissolved in 50% trifluoroaceticacid-CH₂Cl₂ (2 mL) and stirred for 24 h at room temperature. Thevolatiles were removed and the resultant residue was purified by silicagel chromatography to afford the title compound (0.064 g) as a foam(714.2, M+1).

The following compound was prepared by procedures analogous to thosedescribed for the above example:

(R)-2-({(S)-1-[(R)-7-(3,5-Dichloro-phenyl)-5-methyl-6-oxo-5-(4-pyrimidin-5-yl-benzyl)-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carbonyl}-amino)-3-hydroxy-propionicacid (714.6, M+1)

Example 21 Synthesis of(S)-1-[(R)-5-[4-(4-amino-pyrimidin-5-yl)-benzyl]-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-hydroxy-2-methyl-propyl)-amide:

To a stirred solution of(R)-5-(4-bromo-benzyl)-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonylchloride (see Example 14) (1.5 g) in a mixture of CH₂Cl₂ (25 mL) andanhydrous DMF (5 mL) was added L-Pyrrolidine-2-carboxylic acid(2-hydroxy-2-methyl-propyl)-amide (0.84 g) followed byN,N-diisopropylethylamine (1.5 mL). The reaction was stirred at roomtemperature for 2 h then diluted with EtOAc, washed with 1N HCl, H₂O andbrine. The combined organic layers were dried (MgSO₄), filtered andconcentrated. The residue was purified by silica gel chromatography toafford the desired product (1.58 g).

To a stirred solution of the above amide (1.2 g) in DMF (80 mL) wasadded bis(pinacolato)diboron (0.871 g) followed by potassium acetate(0.674 g). The mixture was degassed for 10 min, then PdCl₂(dppf).CH₂Cl₂(0.140 g) was added and the reaction mixture was heated at 80° C. After36 h, the mixture was cooled to room temperature, diluted with H₂O, thenextracted with EtOAc. The combined organic layers were washed with H₂Oand brine, then dried (MgSO₄), filtered and concentrated. The residuewas purified by silica gel chromatography to afford the desired boronate(0.586 g).

The above boronate (0.2 g) was dissolved in DME:H₂O (5 mL: 1 mL).2-Amino-3-bromo-pyrimidine (0.070 g) and potassium carbonate (0.092 g)were added and the flask was flushed with N₂. The mixture was stirredfor 20 min, and then PdCl₂(dppf).CH₂Cl₂ (0.020 g) was added. The mixturewas heated at 80° C. for 2 h, cooled to room temperature and dilutedwith EtOAc. The mixture was filtered, concentrated and the residue waspurified by silica gel chromatography to afford the title compound(0.080 g) as a foam (713.3, M+1)

The following compounds were prepared by procedures analogous to thosedescribed for the above example:

(S)-1-{(R)-7-(3,5-Dichloro-phenyl)-5-[4-(2-fluoro-pyrimidin-5-yl)-benzyl]-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl}-pyrrolidine-2-carboxylicacid (2-hydroxy-2-methyl-propyl)-amide (716.3, M+1)

(S)-1-[(R)-5-[4-(4-Amino-pyrimidin-5-yl)-benzyl]-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-hydroxy-ethyl)-amide (685.1, M+1)

(S)-1-{(R)-7-(3,5-Dichloro-phenyl)-5-[4-(2-fluoro-pyrimidin-5-yl)-benzyl]-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl}-pyrrolidine-2-carboxylicacid (2-hydroxy-ethyl)-amide (688.0, M+1)

(S)-1-[(R)-5-[4-(2-Cyano-pyridin-3-yl)-benzyl]-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid (2-hydroxy-2-methyl-propyl)-amide (722.1, M+1)

(S)-1-[(R)-5-[4-(2-Cyano-pyridin-3-yl)-benzyl]-7-(3,5-dichloro-phenyl)-5-methyl-6-oxo-6,7-dihydro-5H-imidazo[1,2-a]imidazole-3-sulfonyl]-pyrrolidine-2-carboxylicacid carbamoylmethyl-amide (707.1, M+1)

Description of Biological Properties

The biological properties of representative compounds of the formula Iwere investigated by way of the experimental protocol described below.

Assay to Determine Inhibition of LFA-1 Binding to ICAM-1

Purpose of Assay:

This assay protocol is designed to study the direct antagonism, by atest compound, of the interaction of the CAM, ICAM-1 with theLeukointegrin CD18/CD11a (LFA-1).

Description of Assay Protocol:

LFA-1 is immunopurified using the TS2/4 antibody from a 20 g pellet ofhuman JY or SKW3 cells, utilizing a protocol previously described(Dustin, M. J.; et al., J. Immunol. 1992, 148, 2654-2660). The LFA-1 ispurified from SKW3 lysates by immunoaffinity chromatography on TS2/4LFA-1 mAb Sepharose and eluted at pH 11.5 in the presence of 2 mM MgCl₂and 1% octylglucoside. After collection and neutralization of fractionsfrom the TS2/4 column, samples are pooled and precleared with Protein Gagarose.

A soluble form of ICAM-1 is constructed, expressed, purified andcharacterized as previously described (Marlin, S.; et al., Nature, 1990,344, 70-72 and see Arruda, A.; et al., Antimicrob. Agents Chemother.1992, 36, 1186-1192). Briefly, isoleucine 454 which is located at theputative boundary between domain 5 of the ectodomain and thetransmembrane domain, is changed to a stop codon using standardoligonucleotide-directed mutagenesis. This construction yields amolecule identical with the first 453 amino acids of membrane boundICAM-1. An expression vector is created with a hamster dihydrofolatereductase gene, a neomycin-resistance marker, and the coding region ofthe sICAM-1 construct described above, along with the promoter, splicesignals, and polyadenylation signal of the SV40 early region. Therecombinant plasmid is transfected into CHO DUX cells using standardcalcium phosphate methods. Cells are passaged in selective media (G418)and colonies secreting sICAM-1 are amplified using methotrexate. sICAM-1is purified from serum-free media using traditional non-affinitychromatographic techniques, including ion exchange and size exclusionchromatography.

LFA-1 binding to ICAM-1 is monitored by first incubating sICAM-1 at 40μg/mL in Dulbecco's phosphate buffered saline with calcium andmagnesium, additional 2 mM MgCl₂ and 0.1 mM PMSF (Diluting Buffer) in a96-well plate for 30 min at room temperature. Plates are then blocked bythe addition of 2% (w/v) bovine serum albumin in Diluting Buffer for 37°C. for 1 h. Blocking solution is removed from wells, and test compoundsare diluted and then added followed by the addition of approximately 25ng of immunoaffinity purified LFA-1. The LFA-1 is incubated in thepresence of test compound and ICAM-1 at 37° C. for 1 h. Wells are washed3 times with Diluting Buffer. The bound LFA-1 is detected by theaddition of a polyclonal antibody directed against a peptidecorresponding to the CD 18 cytoplasmic tail in a 1:100 dilution withDiluting Buffer and 1% BSA and allowed to incubate for 45 min at 37° C.Wells are washed 3 times with Diluting Buffer and the bound polyclonalantibody is detected by the addition of a 1:4000 dilution of horseradish peroxidase conjugated to goat immunoglobulin directed againstrabbit immunoglobulin. This reagent is allowed to incubate for 20 min at37° C., wells are washed as above and the substrate for the horse radishperoxidase is added to each well to develop a quantitative colorimetricsignal proportional to the amount of LFA-1 bound to sICAM-1. SolubleICAM-1 (60 μg/mL) is used as a positive control for inhibition of theLFA-1/ICAM-1 interaction. The lack of the addition of LFA-1 to thebinding assay is used as a background control for all samples. Adose-response curve is obtained for all test compounds.

All compounds made in the above examples were tested in this assay andeach found to have a K_(d)<10 μM.

Assay to Determine Metabolism by Human Liver Microsomal Enzymes

Purpose of Assay:

This assay protocol is designed to measure the in vitro metabolism oftest compounds by human liver microsomal enzymes. The data collected areanalyzed to calculate a half-life (t_(1/2), min) for test compounds.

Description of Assay Protocol:

The assay is performed in 50 mM potassium phosphate buffer, pH 7.4 and2.5 mM NADPH. Test samples are dissolved in acetonitrile for a finalassay concentration of 1-10 μM. Human liver microsomes are diluted inassay buffer to a final assay concentration of 1 mg protein/mL. A volumeof 25 μL compound solution and 50 μL microsome suspension are added to825 μL assay buffer. The preparation is incubated for 5 min in a 37° C.water bath. The reaction is started by the addition of 100 μL NADPH.Volumes of 80 μL are removed from the incubation mix at 0, 3, 6, 10, 15,20, 40, and 60 min after the start of the reaction and added to 160 μLacetonitrile. The samples are shaken for 20 sec and then centrifuged for3 min at 3000 rpm. A 200 μL volume of the supernatant is transferred to0.25 mm glass fiber filter plates and centrifuged for 5 min at 3000 rpm.Injection volumes of 10 μL are typically added to Zorbax SB C8 HPLCcolumns with formic acid in water or acetonitrile at a flow rate of 1.5mL/min. Percent loss of parent compound is calculated from the areaunder each time point to determine the half-life.

Compounds made in the above examples were tested in this assay andgenerally found to have a t_(1/2)≧40 minutes.

Description of Therapeutic Use

The novel small molecules of formula I provided by the invention inhibitthe ICAM-1/LFA-1 dependent homotypic aggregation of human lymphocytesand human lymphocyte adherence to ICAM-1. These compounds havetherapeutic utility in the modulation of immune cellactivation/proliferation, e.g., as competitive inhibitors ofintercellular ligand/receptor binding reactions involving CAMs andLeukointegrins. To be more specific, the compounds of the invention maybe used to treat certain inflammatory conditions, including conditionsresulting from a response of the non-specific immune system in a mammal(e.g., adult respiratory distress syndrome, shock, oxygen toxicity,multiple organ injury syndrome secondary to septicemia, multiple organinjury syndrome secondary to trauma, reperfusion injury of tissue due tocardiopulmonary bypass, myocardial infarction or use with thrombolysisagents, acute glomerulonephritis, vasculitis, reactive arthritis,dermatosis with acute inflammatory components, stroke, thermal injury,hemodialysis, leukapheresis, ulcerative colitis, necrotizingenterocolitis and granulocyte transfusion associated syndrome) andconditions resulting from a response of the specific immune system in amammal (e.g., psoriasis, organ/tissue transplant rejection, graft vs.host reactions and autoimmune diseases including Raynaud's syndrome,autoimmune thyroiditis, dermatitis, multiple sclerosis, rheumatoidarthritis, insulin-dependent diabetes mellitus, uveitis, inflammatorybowel disease including Crohn's disease and ulcerative colitis, andsystemic lupus erythematosus). The compounds of the invention may alsobe used in treating asthma or as an adjunct to minimize toxicity withcytokine therapy in the treatment of cancers. In general these compoundsmay be employed in the treatment of those diseases currently treatablethrough steroid therapy.

Thus, another aspect of the invention is the provision of a method forthe treatment or prophylaxis of the above-described conditions throughthe adminstration of therapeutic or prophylactic amounts of one or morecompounds of the formula I.

In accordance with the method provided by the invention, the novelcompounds of formula I may be administered for either a prophylactic ortherapeutic purpose either alone or with other immunosuppressive orantiinflammatory agents. When provided prophylactically, theimmunosuppressive compound(s) are provided in advance of anyinflammatory response or symptom (for example, prior to, at, or shortlyafter the time of an organ or tissue transplant but in advance of anysymptoms of organ rejection). The prophylactic administration of acompound of the formula I serves to prevent or attenuate any subsequentinflammatory response (such as, for example, rejection of a transplantedorgan or tissue, etc.). The therapeutic administration of a compound ofthe formula I serves to attenuate any actual inflammation (such as, forexample, the rejection of a transplanted organ or tissue). Thus, inaccordance with the invention, a compound of the formula I can beadministered either prior to the onset of inflammation (so as tosuppress an anticipated inflammation) or after the initiation ofinflammation.

The novel compounds of the formula I may, in accordance with theinvention, be administered in single or divided doses by the oral,parenteral or topical routes. A suitable oral dosage for a compound offormula I would be in the range of about 0.1 mg to 10 g per day. Inparenteral formulations, a suitable dosage unit may contain from 0.1 to250 mg of said compounds, whereas for topical administration,formulations containing 0.01 to 1% active ingredient are preferred. Itshould be understood, however, that the dosage administration frompatient to patient will vary and the dosage for any particular patientwill depend upon the clinician's judgement, who will use as criteria forfixing a proper dosage the size and condition of the patient as well asthe patient's response to the drug.

When the compounds of the present invention are to be administered bythe oral route, they may be administered as medicaments in the form ofpharmaceutical preparations which contain them in association with acompatible pharmaceutical carrier material. Such carrier material can bean inert organic or inorganic carrier material suitable for oraladministration. Examples of such carrier materials are water, gelatin,talc, starch, magnesium stearate, gum arabic, vegetable oils,polyalkylene-glycols, petroleum jelly and the like.

The pharmaceutical preparations can be prepared in a conventional mannerand finished dosage forms can be solid dosage forms, for example,tablets, dragees, capsules, and the like, or liquid dosage forms, forexample solutions, suspensions, emulsions and the like. Thepharmaceutical preparations may be subjected to conventionalpharmaceutical operations such as sterilization. Further, thepharmaceutical preparations may contain conventional adjuvants such aspreservatives, stabilizers, emulsifiers, flavor-improvers, wettingagents, buffers, salts for varying the osmotic pressure and the like.Solid carrier material which can be used include, for example, starch,lactose, mannitol, methyl cellulose, microcrystalline cellulose, talc,silica, dibasic calcium phosphate, and high molecular weight polymers(such as polyethylene glycol).

For parenteral use, a compound of formula I can be administered in anaqueous or non-aqueous solution, suspension or emulsion in apharmaceutically acceptable oil or a mixture of liquids, which maycontain bacteriostatic agents, antioxidants, preservatives, buffers orother solutes to render the solution isotonic with the blood, thickeningagents, suspending agents or other pharmaceutically acceptableadditives. Additives of this type include, for example, tartrate,citrate and acetate buffers, ethanol, propylene glycol, polyethyleneglycol, complex formers (such as EDTA), antioxidants (such as sodiumbisulfite, sodium metabisulfite, and ascorbic acid), high molecularweight polymers (such as liquid polyethylene oxides) for viscosityregulation and polyethylene derivatives of sorbitol anhydrides.Preservatives may also be added if necessary, such as benzoic acid,methyl or propyl paraben, benzalkonium chloride and other quaternaryammonium compounds.

The compounds of this invention may also be administered as solutionsfor nasal application and may contain in addition to the compounds ofthis invention suitable buffers, tonicity adjusters, microbialpreservatives, antioxidants and viscosity-increasing agents in anaqueous vehicle. Examples of agents used to increase viscosity arepolyvinyl alcohol, cellulose derivatives, polyvinylpyrrolidone,polysorbates or glycerin. Microbial preservatives added may includebenzalkonium chloride, thimerosal, chloro-butanol or phenylethylalcohol.

Additionally, the compounds provided by the invention can beadministered topically or by suppository.

Formulations

Compounds of the formula I can be formulated for therapeuticadministration in a number of ways. Descriptions of several exemplaryformulations are given below.

Example A

Capsules or Tablets Example A-1 Example A-2 Ingredients QuantityIngredients Quantity Compound of 250 mg Compound of formula I  50 mgformula I Starch 160 mg Dicalcium Phosphate 160 mg Microcrys. Cellulose 90 mg Microcrys. Cellulose  90 mg Sodium Starch  10 mg Stearic acid  5mg Glycolate Magnesium Stearate  2 mg Sodium Starch Glycolate  10 mgFumed colloidal silica  1 mg Fumed colloidal silica  1 mg

The compound of formula I is blended into a powder mixture with thepremixed excipient materials as identified above with the exception ofthe lubricant. The lubricant is then blended in and the resulting blendcompressed into tablets or filled into hard gelatin capsules.

Example B

Parenteral Solutions Ingredients Quantity Compound of formula I 500 mgPEG 400 40% by volume Ethyl Alcohol 5% by volume Saline 55% by volume

The excipient materials are mixed and then added to one of the compoundsof formula I in such volume as is necessary for dissolution. Mixing iscontinued until the solution is clear. The solution then filtered intothe appropriate vials or ampoules and sterilized by autoclaving.

Example C

Suspension Ingredients Quantity Compound of formula I 100 mg Citric acid1.92 g Benzalkonium chloride 0.025% by weight EDTA 0.1% by weightPolyvinylalcohol 10% by weight Water q.s. to 100 mL

The excipient materials are mixed with the water and thereafter one ofthe compounds of formula I is added and mixing is continued until thesuspension is homogeneous. The suspension is then transferred into theappropriate vials or ampoules.

Example D

Topical Formulation Ingredients Quantity Compound of formula I   5% byweight Tefose 63   13% by weight Labrafil M 1944 CS   3% by weightParaffin Oil   8% by weight Methylparaben (MP) 0.15% by weightPropylparaben (PP) 0.05% by weight Deionized water q.s. to 100

The proper amounts of Tefose 63, Labrafil M 1944 CS, Paraffin oil andwater are mixed and heated at 75° C. until all components have melted.The mixture is then cooled to 50° C. with continuous stirring.Methylparaben and propylparaben are added with mixing and the mixture iscooled to ambient temperature. The compound of formula I is added to themixture and blended well.

1. A method for treating inflammation or an inflammatory condition in apatient which comprises administering to said patient a therapeuticallyeffective amount of a compound a compound of the formula I:

wherein: R¹ and R² are each independently selected from the groupconsisting of: (A) hydrogen, with the proviso that R¹ and R² are notboth hydrogen atoms; (B) —R¹⁰⁰, which is:  a straight or branched alkylof 1 to 7 carbon atoms or cycloalkyl of 3 to 6 carbon atoms, which alkylor cycloalkyl group is mono- or poly substituted with moietiesindependently selected from the group consisting of: (i) oxo, (ii)cyano, (iii) halogen, (iv) moieties of the formula —COOR⁶, wherein R⁶ isa hydrogen atom, a straight or branched alkyl of 1 to 7 carbon atoms orcycloalkyl of 3 to 6 carbon atoms, (v) moieties of the formula —OR,wherein R is a hydrogen atom, a straight or branched alkyl group of 1 to7 carbon atoms or an acyl group of the formula —COR⁸ wherein R⁸ is astraight or branched alkyl group of 1 to 7 carbon atoms, (vi) moietiesof the formula —NR⁹R¹⁰, wherein R⁹ and R¹⁰ are each, independentlyselected from the group consisting of: (a) hydrogen, (b) straight orbranched alkyl of 1 to 7 carbon atoms, (c) acyl of the formula —COR¹¹wherein R¹¹ is a straight or branched alkyl group of 1 to 7 carbonatoms, and (d) groups of the formula —COOR¹² wherein R¹² is a straightor branched alkyl group of 1 to 7 carbon atoms,  or wherein R⁹ and R¹⁰constitute a bridge consisting of 3-5 methylene groups or 2-4 methylenegroups and one oxygen atom, such that the groups R⁹ and R¹⁰ togetherwith the nitrogen atom between them form a heterocyclic ring, (vii)saturated heterocyclic groups, consisting of 3 to 5 methylene groups andone oxygen atom, wherein said heterocyclic groups are optionally mono-or disubstituted with moieties that are independently selected from thegroup consisting of: (a) oxo and (b) straight or branched alkyl of 1 to3 carbon atoms; and (viii) aryl, selected from the group consisting of:(a) furyl, (b) tetrazolyl and (c) thiophenyl; (C) aryl, selected formthe group consisting of: (i) biphenyl, (ii) phenyl which is mono- ordi-substituted with moieties independently selected from the groupconsisting of —NH₂ and N-morpholino, and (iii) quinolinyl; and (D)unsaturated or partially saturated heterocyclic groups consisting of 2to 3 carbon atoms, 1 to 2 nitrogen atoms, 0 to 1 sulfur atoms and 0 to 1oxygen atoms wherein said heterocyclic group is optionally mono- orpolysubstituted with one or more of the following moieties independentlyselected from the group consisting of: (i) oxo and (ii) straight orbranched alkyl of 1 to 7 carbon atoms;  or wherein R¹ and R² constitutea saturated 3 to 5 methylene group bridge which together with thenitrogen atom between them form a heterocyclic ring, wherein saidheterocyclic ring is mono- or disubstituted with moieties independentlyselected from the group consisting of: (A) —OH, (B) —COOH and (C)—CONH₂; R³ is: (A) aryl selected from the group consisting of pyridyland pyrimidyl, wherein one or more hydrogen atoms of said aryl group areoptionally and independently substituted with moieties selected from thegroup consisting of: (i) cyano, (ii) halogen and (iii) groups of theformula —NR¹³R¹⁴, wherein R¹³ and R¹⁴ are each, independently, hydrogenor straight or branched alkyl of 1 to 3 carbon atoms; (B)trifluoromethoxy or, (C) cyano; R⁴ is straight or branched alkyl of 1 to3 carbon atoms; R^(5a) is C₁ or CF₃; R^(5b) is C₁ or CF₃; X is an oxygenor a sulfur atom; and Y is an oxygen or a sulfur atom; or apharmaceutically acceptable salt thereof.
 2. The method of claim 1wherein the condition to be treated is adult respiratory distresssyndrome, shock, oxygen toxicity, multiple organ injury syndromesecondary to septicemia, multiple organ injury syndrome secondary totrauma, reperfusion injury of tissue due to cardiopulmonary bypass,myocardial infarction or use with thrombolysis agents, acuteglomerulonephritis, vasculitis, reactive arthritis, dermatosis withacute inflammatory components, stroke, thermal injury, hemodialysis,leukapheresis, ulcerative colitis, necrotizing enterocolitis orgranulocyte transfusion associated syndrome.
 3. The method of claim 1wherein the condition to be treated is psoriasis, organ/tissuetransplant rejection, graft vs. host reaction or an autoimmune disease.4. The method of claim 1 wherein the condition to be treated ispsoriasis.
 5. The method of claim 1 wherein the condition to be treatedis Raynaud's syndrome, autoimmune thyroiditis, dermatitis, multiplesclerosis, rheumatoid arthritis, insulin-dependent diabetes mellitus,uveitis, inflammatory bowel disease, Crohn's disease, ulcerative colitisor systemic lupus erythematosus.
 6. The method of claim 1 wherein thecondition to be treated is multiple sclerosis.
 7. The method of claim 1wherein the condition to be treated is asthma.
 8. The method of claim 1wherein the condition to be treated is the toxic effects of cytokinetherapy.